Investigation into the mechanism of replication of inc P-1 and inc P-4 broad host-range plasmids will be conducted. Emphasis will be placed on cloning the genetic determinants of P-1 incompatibility, obtaining mutations in this region as a means of analyzing function, and identifying physically the protein products involved. Plasmid genes essential for replication of the inc P-4 plasmid R1162 will be identified and mapped. Work will be initiated on an in vitro system of DNA replication for this plasmid. Transposable derivatives of broad host-range plasmids will be constructed by replacing the central Hind III fragment of the transposon Tn 5 with plasmid DNA. This will enable the insertion of broad host-range replicons into the other plasmids, possibly increasing the host-range of the latter.